The Ras/MAP kinase signaling pathway promotes cell divisions during the development of many multicellular organisms. Study of this signaling pathway is significant since mutation of the ras gene in humans is an important step in the generation of cancerous tumors. Many proteins are involved in the regulation and transmission of the Ras/MAP kinase signal, and we have initiated a study of a novel target of this signaling pathway during Caenorhabditis elegans development. The C. elegans predicted open-reading-frame, T08D10.1, codes for member of the CCAAT-box family of DNA-binding proteins and possesses multiple potential phosphorylation sites and docking sites for extracellular regulated kinase (ERK) MAP kinase. Preliminary data indicate that T08D10.1 acts as a high affinity substrate for ERK MAP kinase and functions to inhibit Ras signaling during C. elegans vulval development. We propose to characterize the function and expression of this novel component of the Ras signaling pathway through biochemical and genetic methods. The ability of T08D10.1 to function as a substrate for ERK MAP kinase and the role of putative docking site sequences will be investigated using biochemical kinase assays. The mutant phenotype of T08D10.1 will be characterized during C. elegans development in wild-type and sensitizes backgrounds. The temporal and spatial expression pattern of T08D10.1 will be investigated through the generation of transgenic animals. Finally, the function of T08D10.1 as a transcription factor will be tested using in vitro electrophoretic mobility shift assays and transactivation potential assays conducted in cultured cells. Investigation of the role of T08D10.1 as a downstream mediator of the Ras pathway will not only contribute to the understanding of the mechanisms required for the specification of specific cell fates during the development of C. elegans, but will also contribute to our understanding of the general mechanisms by which general uptstream signaling events lead to appropriate cell fate decisions during normal and abnormal metazoan development. Mutation of components of the Ras signaling pathway can lead to the development of cancer in humans. Many of the core components of this pathway are conserved in multicellular organisms including the model organism Caenorhabditis elegans. This project seeks to characterize a novel regulator of the Ras signaling pathway in C. elegans that may function to inhibit cell divisions. [unreadable] [unreadable] [unreadable]